Modulation by cAMP of 1alpha,25-dihydroxyvitamin D3 sensitivity of murine erythroleukemia cells.

As we previously reported, 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) dose-dependently inhibited not only proliferation of undifferentiated murine erythroleukemia (MEL) cells but also activin A-induced erythroid differentiation of MEL cells. However, the effect of 1,25(OH)2D3 on MEL cell proliferation was significantly greater by one order of magnitude than that on differentiation (IC(50): 9.2 vs 0.8 nM, respectively). The response of activin A-treated mature MEL cells to 1,25(OH)2D3 in the induction of 1,25(OH)2D3-24-hydroxylase (24-OHase) activity, a rapid effect of 1,25(OH)2D3, was enhanced to the same degree as in untreated immature cells, suggesting that differences in capacity of cells to inactivate 1,25(OH)2D3 did not contribute to augmentation of 1,25(OH)2D3 effect in activin A-treated mature cells. Furthermore, neither the number nor the affinity of vitamin D receptors (VDR) differed significantly between activin A-treated cells and untreated immature cells. The intracellular cAMP level, which affects 1,25(OH)2D3-mediated induction of 24-OHase activity, was significantly less in activin A-treated mature cells than in immature MEL cells. The addition of dibutyryl cAMP (dbc AMP) to activin A-treated MEL cells dose-dependently attenuated 1,25(OH)2D3-mediated induction of 24-OHase activity, finally to a level comparable to that of the untreated cells at the final concentration of 100 nM dbcAMP, while dbcAMP itself by 100 nM did not affect MEL cell differentiation by 24 h. In summary, we have shown for the first time that 1,25(OH)2D3 exerted its effect on leukemia cells at physiological concentration and that the magnitude of this effect depended on the changes in intracellular cAMP level through stages of differentiation, suggesting that the cAMP-protein kinase A system may be useful as a target for clinical application of vitamin D analogs by improving the sensitivity of leukemic cells to 1,25(OH)2D3.

Mechanism of atherosclerotic calcification.

Calcification is almost invariably associated with atherosclerotic plaque lesions. Recent data suggest that plaque calcification is an active, regulated process similar to osteogenesis. In order to clarify the mechanism of plaque calcification, we developed an in vitro model of vascular calcification by utilizing bovine vascular smooth muscle cells (BVSMCs). This model is useful in that diffuse and massive calcification can be induced within 2 weeks and thereby biochemical analyses of vascular calcification can be performed. We have analyzed several aspects of vascular calcification by using this model and demonstrated as follows: 1) in vitro calcification of BVSMCs is regulated by calciotropic hormones and BVSMCs are equipped with a unique autocrine and/or paracrine system regulating calcium metabolism. 2) Sodium-dependent phosphate cotransport plays a crucial role in BVSMC calcification as well as in mineralization of skeletal tissues. 3) BVSMCs acquire osteoblastic phenotype under certain conditions. Finally, we discuss the roles of macrophages in the development of atherosclerotic calcification. Interferon-gamma (IFN-gamma) induces gene expression of 25-hydrovitamin D-1 alpha-hydroxylase (1 alpha OHase) and its activity in macrophages. Since 1 alpha OHase can locally convert 25-hydroxyvitamin D into 1 alpha, 25-dihydroxyvitamin D (1,25(OH)2D), an active metabolite of vitamin D, it is suggested that local production of 1,25(OH)2D by macrophages may promote atherosclerotic calcification. Moreover, macrophages may be involved in the phenotypic changes of vascular smooth muscle cells (VSMCs) to acquire calcifying capacity. Therefore, the phenotypic changes of VSMCs in atherosclerotic plaque may contribute to the development of atherosclerotic calcification.

Synergistic enhancement by 12-O-tetradecanoylphorbol-13-acetate and dibutyryl cAMP of 1alpha,25-dihydroxyvitamin D3 action in human promyelocytic leukemic HL-60 cells.

We have reported that dibutyryl cAMP (dbcAMP), an activator of cAMP-dependent protein kinase (PKA), potentiated the effects of 1alpha,25-dihydroxyvitamin D3(1,25-(OH)2D3)-induced 24-hydroxylation activity in HL-60 cells by increasing 1,25-(OH)2D3 receptor (VDR). The present study demonstrated that 12-O-tetradecanoylphorbol-13-acetate (TPA), a potent phorbol ester, also potentiated the effect of 1,25-(OH)2D3 on HL-60 cells and that TPA and dbcAMP acted in a synergistic manner to enhance the effect of 1,25-(OH)2D3. It is interesting that TPA induced 24-hydroxylation activity far more efficiently than dbcAMP, in addition to their effects in increasing VDR. TPA increased basal levels of c-fos mRNA to the maximum by 1 h after the treatment, whereas dbcAMP failed to affect c-fos gene expression. Together with the previous data indicating the presence of AP-1-like sequence in the promoter of 24-hydroxylase gene, it was suggested that TPA potentiated the effect of 1,25-(OH)2D3 through an activation of c-fos gene expression. This notion was further supported by the data showing that TPA and dbcAMP also acted in a synergistic manner to activate c-fos gene expression. Neither TPA nor dbcAMP affected c-jun early response gene in the HL-60 clone used in the present study. The present study suggested that the activation of early c-fos response gene by TPA might be another mechanism to enhance the effect of 1,25-(OH)2D3, besides up-regulation of VDR.

Homeostasis model assessment as a clinical index of insulin resistance in type 2 diabetic patients treated with sulfonylureas.

OBJECTIVE: To investigate whether the insulin resistance index (IR) assessed by homeostasis model assessment (HOMA) is associated with the insulin resistance index assessed by euglycemic-hyperinsulinemic clamp (clamp IR) in type 2 diabetic patients who received sulfonylureas (SUs), as well as in those treated by diet alone. RESEARCH DESIGN AND METHODS: Retrospectively, the association between HOMA IR and clamp IR was analyzed in 80 type 2 diabetic subjects (53 subjects treated with SUs and 27 subjects treated with diet alone). The 80 subjects, selected because they had not received insulin therapy, were among 111 diabetic participants in a clamp study for evaluation of insulin resistance from May 1993 to December 1997 in Osaka City University Hospital. RESULTS: The HOMA IR showed a hyperbolic relationship with clamp IR. The log-transformed HOMA IR (all subjects, r = -0.725, P < 0.0001; SU group, r = -0.727, P < 0.0001; diet group, r = -0.747, P < 0.0001) correlated more strongly with clamp IR than did HOMA IR per se (all subjects, r = -0.594, P < 0.0001; SU group, r = -0.640, P < 0.0001; diet group, r = -0.632, P = 0.0004). The univariate regression line between log-transformed HOMA IR and clamp IR in the SU group did not differ from that in the diet group (slope, -6.866 vs. -5.120, P > 0.05; intercept, 6.566 vs. 5.478, P > 0.05). Stepwise multiple regression analyses demonstrated that the log-transformed HOMA IR was the strongest independent contributor to clamp IR (R2 = 0.640, P < 0.0001). CONCLUSIONS: The HOMA IR strongly correlated with the clamp IR in type 2 diabetic patients treated with SUs as well as in those treated with diet alone.

Stiffness indexes beta of the common carotid and femoral arteries are associated with insulin resistance in NIDDM.

OBJECTIVE: To investigate the association between arterial wall stiffness indexes beta of the common carotid artery (CCA) and the femoral artery (FA) and insulin resistance in NIDDM subjects in a cross-sectional study. RESEARCH DESIGN AND METHODS: We evaluated the arterial stiffness indexes beta of CCA and FA using an ultrasonic phase-locked echo-tracking system in 60 NIDDM subjects attending the diabetes center in Osaka City University Hospital, compared with 120 age- and sex-matched control subjects. Insulin sensitivity indexes were evaluated using a euglycemic-hyperinsulinemic clamp. RESULTS: Stiffness indexes beta of both CCA and FA were significantly higher in NIDDM subjects than in control subjects (CCA 18.1 +/- 0.9 vs. 11.7 +/- 0.3, respectively, P < 0.001; FA 35.7 +/- 2.3 vs. 23.7 +/- 0.8, respectively, P < 0.001). The mean insulin sensitivity index in NIDDM subjects was 4.69 +/- 0.29 mg.kg-1.min-1.mU-1.l. The stiffness indexes beta of both CCA and FA were inversely correlated with insulin sensitivity indexes (CCA r = -0.393, P = 0.002; FA r = -0.329, P = 0.010), as well as with age, duration of diabetes, and mean blood pressure. In stepwise multiple regression analyses, insulin sensitivity index and duration of diabetes were identified as significant independent variables for stiffness indexes beta in both CCA and FA (CCA R2 = 0.249, P = 0.0003; FA R2 = 0.336, P < 0.001). CONCLUSIONS: Arterial stiffness indexes beta of CCA and FA were associated with insulin resistance in NIDDM subjects.

Seminal tract obstruction caused by childhood inguinal herniorrhaphy: results of microsurgical reanastomosis.

PURPOSE: Childhood inguinal herniorrhaphy is a frequent causes of seminal tract obstruction. We investigate the characteristics of this obstruction, surgical techniques for reanastomosis and outcomes in patients with bilateral or unilateral obstruction caused by inguinal herniorrhaphy in childhood. MATERIALS AND METHODS: We treated 22 men an average of 30.5 years old in whom average duration of obstruction was 27.1 years. Obstruction was bilateral in 9 patients and unilateral in 13, and 14 had azoospermia preoperatively. Microsurgical 2 layer vasovasostomy and/or specific tubule epididymovasostomy was performed. RESULTS: Inguinal and crossed vasovasostomy was done in 18 and 4 patients, respectively. The distal end of the vas was found at the internal inguinal ring or in the pelvic cavity in 57.1% of the vasa and more than 3 cm. of the vas had been resected in 37%. Sperm was noted in vasal fluid in 45.2% of the vasa during the operation and the patency rate of inguinal vasovasostomy was 88.9%. Ipsilateral epididymovasostomy performed after patent inguinal vasovasostomy in 5 patients with secondary epididymal obstruction resulted in normal sperm density and 3 pregnancies. In 7 men more than 2 procedures were done. There was sperm in the ejaculate in 12 of the 14 patients who had had azoospermia preoperatively and apparently increased sperm density postoperatively in 4 of the 8 who had not had azoospermia preoperatively. Pregnancy was achieved by 7 of the 21 married men (33.3%). CONCLUSIONS: Microsurgical reanastomosis of the seminal tract resulted in high patency and pregnancy rates in cases of seminal tract obstruction caused by childhood inguinal herniorrhaphy. Patients should elect seminal tract reanastomosis or assisted reproductive technology using epididymal or testicular sperm after receiving sufficient information on each treatment modality.

Endoscopic harvesting of the gracilis muscle for reinnervated free-muscle transfer.

Reinnervated functioning free-muscle transfer has proven to be invaluable in numerous reconstructive procedures. However, one problem that remains unsolved after transferring the muscle is the presence of a long and cosmetically unacceptable scar at the donor site. This scar has undermined patients' satisfaction with the procedure despite its excellent functional results. In an attempt to resolve this problem, the authors harvested the gracilis muscle endoscopically and now report their technique and results. To create an optical cavity in harvesting the gracilis muscle endoscopically, they devised a lifting apparatus, which is described. Comparative study showed that endoscopic harvesting of the gracilis produced a significantly shorter scar, but took 1.5 times longer than conventional method.

Insulin resistance in non-obese, non-insulin-dependent diabetic patients with diabetic nephropathy.

To investigate the association between insulin resistance and diabetic nephropathy, peripheral insulin sensitivity indices (M/I values) were evaluated via euglycemic-hyperinsulinemic clamp in 45 non-obese, non-insulin-dependent diabetic (NIDDM) subjects. The patients were divided into four groups: 18 with normoalbuminuria (urinary albumin excretion rate [AER] < 30 mg/24 h, stage I), 10 with microalbuminuria (30 < or = AER < or = 300 mg/24 h, stage II), seven with overt proteinuria (AER > 300 mg/24 h, stage III), and 10 with uremia (serum creatinine levels > 2.0 mg/dL, stage IV). There were no significant differences in age, body mass index (BMI), fasting plasma glucose, or hemoglobin A1c (HbA1c) among the four groups. No significant difference in M/I values was seen between stage I and stage II (6.30 +/- 0.73 and 5.95 +/- 0.85 mg/kg/(min per microU/mL) x 100, respectively). M/I values in the stage I and stage II groups were strongly correlated with BMI (r = -.790, P = .0001 and r = -.785, P = .007, respectively). M/I values in the stage III group (4.53 +/- 0.51) were lower than in the stage I group, although not significantly so. M/I values in the stage IV group (3.16 +/- 0.37) were significantly lower than in the stage I group (P = .025). In multiple regression analysis with a model in which age, sex, BMI, HbA1c, and creatinine clearance (Ccr) were included as independent variables, BMI and Ccr were demonstrated to be significant and independent contributors to insulin sensitivity indices as the dependent variable (beta = -0.716 and beta = 0.272, respectively, R2 = .564, P < .0001). In conclusion, the present cross-sectional study demonstrated in non-obese NIDDM patients with nephropathy that microalbuminuria did not affect peripheral insulin resistance, but uremia did, as in nondiabetic patients, and that the peripheral insulin resistance was significantly contributed to by the degree of obesity and uremia.

Significance of elbow extension in reconstruction of prehension with reinnervated free-muscle transfer following complete brachial plexus avulsion.

Thirty-one patients with complete avulsion of the brachial plexus underwent reconstruction of elbow extension by intercostal nerve transfer following reconstruction of prehension with either a single or double free-muscle transfer. Long-term results of elbow extension were evaluated in 24 patients. Reinnervation of the triceps muscle took longer than that of the transferred muscle on serial electromyographic examinations, and the eventual strength of the triceps muscle was weak. None attained M5 grade, 2 achieved M4 grade, 4 achieved M3 grade, 8 achieved M2 grade, 5 achieved M1 grade, and another 5 achieved M0 grade. However, despite the weak recovery, 14 patients were able to obtain useful functional recovery of the triceps muscle, enabling it to stabilize the elbow joint against the transferred muscle, which acted as simultaneous elbow flexor and wrist or finger extensor. Elbow stability is imperative in order to obtain voluntary finger function following free-muscle transfer. Should the triceps muscle fail to recover following intercostal nerves neurotization, transferring the reinnervated infraspinatus to the triceps is an optional procedure to provide stabilization of the elbow.

Renal function and insulin resistance as determinants of plasma leptin levels in patients with NIDDM.

Plasma leptin level is known to correlate with the degree of obesity. To determine the influences of renal function and insulin resistance on plasma leptin concentrations, we measured plasma leptin concentrations and performed the euglycaemic hyperinsulinaemic clamp studies in 57 patients with non-insulin-dependent diabetes mellitus with a wide range of renal function. In simple regression analyses, plasma leptin concentration showed significant positive correlations with percentage of body fat measured by dual energy X-ray absorptiometry, body mass index, waist to hip ratio and fasting plasma insulin. Leptin level was higher in females than males. Multiple regression analyses indicated that percent body fat, waist to hip ratio, plasma insulin, gender and renal function (1/creatinine), but not insulin sensitivity, were significant and independent determinants of plasma leptin level. These results suggest that plasma leptin level is regulated or affected by multiple factors including renal function. Insulin resistance appeared to increase leptin levels indirectly by raising plasma insulin.

[Clinical statistics on in-patients and operations during a 20-year period (1975-1994) at Department of Urology, Kansai Medical University].

The patients, diseases and operations experienced between 1975 and 1994 in our department were statistically analyzed. The numbers of in-patients and operations have been increasing since 1977. During these 20 years, endoscopic surgery has replaced many open surgical procedures. The introduction of extracorporeal shock wave lithotripsy has dramatically changed the therapeutic modality for urolithiasis, and decreased of the necessity of open surgery.

Quantitative analysis of testicular histology in patients with vas deferens obstruction caused by childhood inguinal herniorrhaphy: comparison to vasectomized men.

PURPOSE: We investigated the effects on spermatogenesis of lifelong vas deferens obstruction caused by childhood inguinal herniorrhaphy. MATERIALS AND METHODS: Testicular histology was analyzed quantitatively in 15 post-herniorrhaphy patients with vasal obstruction for a mean of 28.5 years (group 1). The results were compared to those of 19 vasectomy patients with obstruction for a mean of 8.2 years (group 2). RESULTS: There were significantly fewer total germ cells per tubular cross section in group 1 than in group 2 (p < 0.05). There was a significant negative correlation between the duration of obstruction and total germ cells per tubule (r = -0.389, p < 0.03, 34 patients). The tubular diameter was significantly larger in group 2 than in group 1 (p < 0.01). CONCLUSIONS: Patients with vas deferens obstruction caused by childhood inguinal herniorrhaphy have decreased spermatogenesis compared to post-vasectomy patients, probably due to the longer obstruction period.

Amyloid goiter: radiological study in a case presenting hypothyroidism.

We report herein a case of amyloid goiter associated with rheumatoid arthritis in which hypothyroidism was observed. A 52-year-old housewife who had suffeed from rheumatoid arthritis for 15 years was referred to our hospital because of general fatigue. On admission, a large goiter was observed. Laboratory data showed primary hypothyroidism. Renal biopsy and gastric mucosa biopsy showed amyloid deposition of AA-type. Thyroid biopsy showed massive amyloid involvement. Although the findings of iodine-123 scintigraphy, technetium-99m pertechnetate scintigraphy, computed tomography and magnetic resonance image studies were similar to those for goiter associated with chronic thyroiditis, tallium-201 chloride scintigraphy gave a differing result, demonstrating absent uptake at 3 hours in this case. Replacement therapy with levothyroxine relieved the symptoms. This case was unusual in that amyloid goiter presented clinically as hypothyroidism. Absence of tallium-201 chloride uptake at 3 hours may be a diagnostic specificity for amyloid goiter in differentiating its hypothyroidism from that caused by chronic thyroiditis.

One-stage treatment of infected bone defects of the tibia with skin loss by free vascularized osteocutaneous grafts.

Twenty-six patients who had an infected nonunion or segmental defect of the tibia with skin loss were treated in one stage with debridement and a free vascularized osteocutaneous fibula or iliac graft. Successful control of infection, closure of skin defects, and incorporation of bone union were achieved in all patients except one. In an average follow-up of 39 months, no recurrence of infection was seen. There were four stress fractures of the grafted fibula; these healed within 2 months with cast immobilization. Graft hypertrophy was common in the fibula grafts, but it took 1 1/2 years for hypertrophy of the graft to be strong enough to remove external supports without stress fracture or an additional cancellous bone graft. The use of a one-stage free vascularized osteocutaneous graft for the management of infected bone defects of the tibia with skin loss is effective because extensive debridement can remove all devitalized and infected tissue and can increase vascularity in the region of infection and osseous defect to enhance antibiotic delivery.